Survival outcomes remain poor for lung cancer patients, in part due to genetic alterations-mediated survival signaling pathways in lung cancer cells and treatment resistance. Bax functions as an essential gateway to apoptotic cell death. We previously discovered that the serine (S)184 phosphorylation site of Bax is a critical switch to functionally control Bax's proapoptotic activity. AKT and PKC? have been identified as physiologic Bax kinases that can directly phosphorylate Bax at S184 site leading to inactivation of its proapoptotic function. It is known that genetic mutations in K-Ras, EGFR and PTEN can activate PI3K/AKT survival pathway leading to increased resistance to radiotherapy or chemotherapy in various cancers, including lung cancer. Increased levels of phospho-Bax (i.e. pBax) were observed in tumor tissues in patients with non-small cell lung cancer (NSCLC). We hypothesize that pBax may provide a new predictive and prognostic biomarker in NSCLC. Expression of constitutively active K-Ras G12D mutant or treatment with radiation, cisplatin or rapamycin resulted in activation of AKT and/or PKC? leading to increased phosphorylation of Bax, which may contribute to radio-, chemo- or rapalog resistance. Since increased levels of pBax (i.e. inactive form of Bax) were also observed in both radioresistant and rapalog resistant lung cancer cells as well as K-Ras mutant lung cancer, development of novel small molecules that activate Bax should be critical for treatment of K-Ras mutant lung cancer or overcoming radio- or rapalog resistance. We have identified three small molecule Bax activators (SMBA1-3) that selectively bind the S184 pocket of Bax protein but do not bind other Bcl2 family members. The lead compound SMBA1 not only reverses radioresistance but also overcomes rapalog resistance in vitro. SMBA1 potently represses lung cancer xenografts through induction of apoptosis by activating Bax in tumor tissues. In this proposal, two specific aims have been identified: (1) To determine whether and how genetic alterations or mTOR inhibition regulate Bax phosphorylation leading to Bax inactivation and treatment resistance in human lung cancer cells. Studies will determine whether pBax is a novel prognostic biomarker or therapeutic target in NSCLC patients; (2) Development of novel small molecule Bax activators (SMBA) by targeting the structural pocket around the Bax phosphorylation site for lung cancer therapy. Studies will test the antitumor efficacy of SMBA or in combination with mTOR inhibitor RAD001 or Bcl2 BH4 antagonist BDA-366 in patient-derived xenograft (PDX), radioresistant lung cancer and genetically engineered K-Ras-driven lung cancer animal models. It is expected that a new class of anti-cancer agents and optimal strategies for lung cancer treatment will be developed by targeting Bax.